Zoo-FISH analysis of dog chromosome 5: identification of conserved synteny with human and cat chromosomes

Abstract

Conserved segments of synteny between the human genome and chromosome 5 (CFA 5) of the domestic dog (Canis familiaris) have been identified by reciprocal chromosome painting analysis. A CFA 5 paint probe was applied to human metaphase spreads, revealing distinct hybridisation sites on human (HSA) chromosomes 1, 11, 16, and 17. Paint probes for these human chromosomes were then hybridised to dog metaphase spreads, identifying the regions of CFA 5 with which homology is shared with the corresponding human chromosome. Application of the CFA 5 paint probe to metaphase spreads of the domestic cat (Felis catus, FCA) demonstrated hybridisation to cat chromosomes C1, D1, E1, and E2. Dog PCR primers for type 1 markers known to lie in the corresponding regions of HSA 11, 16, and 17 were used to isolate dog BAC clones representing four genes. Fluorescence in situ hybridisation analysis confirmed their localisation to CFA 5 and suggested that two of the conserved segments lie in opposing orientations on CFA 5, compared to the human chromosome concerned. A third segment appears to lie in the same orientation on both human and dog chromosomes. No suitable gene markers were available for analysis of the fourth segment. The significance of these findings is discussed with reference to current and future dog genome mapping efforts.

Assignment of the canine tectorin alpha gene (TECTA) to CFA5q12→q13 by FISH and confirmation by radiation hybrid mapping

“…Using the MultiMap package, two-point analysis revealed that TECTA is linked to the marker Ren42N13 situated on CFA5 with a Lod score of 19.67. This chromosome assignment of TECTA to CFA5 is in good agreement with known homology between HSA11q and CFA5q (Thomas et al, 1999). The marker Ren42N13 is linked on the RH map of CFA5 proximal to the Thy-1 cell surface antigen gene (THY1) and the CD3E antigen, epsilon polypeptide (TiT3 complex) gene (CD3E) (Breen et al, 2001).…”

Isolation and chromosomal assignment of canine genomic BAC clones representing 25 cancer-related genes

“…Radiolabelled PCR products representing each canine gene marker were used to screen groups of three filters selected from the RPCI-81 canine genomic BAC library (Li et al, 1999) as previously described (Thomas et al, 1999). Clones generating positive hybridisation signals were obtained as bacterial slants from the Human Genome Mapping Project Resource Centre (HGMP-RC, Babraham, Cambs, UK).…”

“…Canine BAC clones were physically mapped by FISH analysis as previously described (Thomas et al, 1999). Briefly, 500 ng of BAC DNA were labelled with either Spectrum Red-, Orange-or Green-dUTP (Vysis), diethylaminomethylcoumarin (DEAC)-5-dUTP (NEN), or biotin-16-dUTP (Boehringer) by nick translation.…”

Chromosome assignment of six dog genes by FISH, and correlation with dog–human Zoo‐FISH data

“…The PCR products obtained from six gene markers were used to screen a dog bacterial artificial chromosome (BAC) library (Li et al . 1999) as previously described (Thomas et al . 1999).…”

“…At least two BAC clones for each gene marker were assigned to the corresponding chromosomal location (Fig. 1) by FISH analysis onto dog metaphase chromosomes (Thomas et al . 1999), using the nomenclature of Breen et al .…”